Environmental DNA from Marine Waters and Substrates: Protocols for Sampling and eDNA Extraction.

Environmental DNA (eDNA) analysis has emerged in recent years as a powerful tool for the detection, monitoring, and characterization of aquatic metazoan communities, including vulnerable species. The rapid rate of adopting the eDNA approach across diverse habitats and taxonomic groups attests to its value for a wide array of investigative goals, from understanding natural or changing biodiversity to informing on conservation efforts at local and global scales. Regardless of research objectives, eDNA workflows commonly include the following essential steps: environmental sample acquisition, processing and preservation of samples, and eDNA extraction, followed by eDNA sequencing library preparation, high-capacity sequencing and sequence data analysis, or other methods of genetic detection. In this chapter, we supply instructional details for the early steps in the workflow to facilitate researchers considering adopting eDNA analysis to address questions in marine environments. Specifically, we detail sampling, preservation, extraction, and quantification protocols for eDNA originating from marine water, shallow substrates, and deeper sediments. eDNA is prone to degradation and loss, and to contamination through improper handling; these factors crucially influence the outcome and validity of an eDNA study. Thus, we also provide guidance on avoiding these pitfalls. Following extraction, purified eDNA is often sequenced on massively parallel sequencing platforms for comprehensive faunal diversity assessment using a metabarcoding or metagenomic approach, or for the detection and quantification of specific taxa by qPCR methods. These components of the workflow are project-specific and thus not included in this chapter. Instead, we briefly touch on the preparation of eDNA libraries and discuss comparisons between sequencing approaches to aid considerations in project design.